simandou project port development area · 2018-11-21 · 1.1.2 lymphatic filariasis ... sop...

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Rio Tinto Iron Ore

Simfer SA Republic of Guinea

March 2012

Authors: Caroline Maxwell, Dr Mark Divall

March 2012, Final Report

Address all correspondence to Dr Mark Divall [email protected]

This report has been prepared by Shape Consulting, with all reasonable skill, care and

diligence within the terms of the contract with the client, and taking account of the resources

devoted to it by agreement with the client. We disclaim any responsibility to the client and

others in respect of any matters outside the scope of the above. This report is confidential to

the client and we accept no responsibility of whatsoever nature to third parties to whom this

report, or any part thereof, is made known. Any such party relies on the report at their own

risk.

© Shape Consulting Limited. 2012. All rights reserved. This report is prepared solely for the benefit of, and use by, SNC-Lavalin Environnement and Rio Tinto Iron Ore Atlantic Limited and Simfer S.A. and may not be sold, reproduced or in any other way copied or transferred by the customer to anyone else, whether in whole or in part. Shape Consulting owns and retains all intellectual property rights in this report including, without limiting the generality of foregoing, all copyrights.

Rio Tinto Iron Ore Atlantic, Simfer SA Entomological Assessment Port Site – Forécariah, Kaback March 2012

Shape Consulting Limited Confidential

Acknowledgements Shape Consulting Limited would like to acknowledge the following individuals for the support

provided in planning and performing this baseline entomological survey in conjunction with

the Rio Tinto, which was carried out in October 2011.

Rio Tinto – Simfer S.A. Catherine Garcia Community relations manager- Port and rail Frédéric Chenais Superintendent communities Dr Kékoura Camara Community liaison officer

SNC Lavalin Environnement Guy Poirier Logistics manager Michel Bureau Logistics manager Richard Fontaine Vice-president special projects Chantal Roy Project manager Claudia Paz-Miller Project Assistant

Flour Mamadu Yoba Barry Driver

National Malaria Control Programme Denka Camara Entomologist Kalil Keita Entomologist

SHAPE Consulting Limited Dr Mark Divall Project Director Caroline Maxwell Entomologist Astrid Knoblauch Epidemiologist Joan Angel Project Administrator



Table of Contents Table of Contents ................................................................................................................... 4 List of Figures ......................................................................................................................... 6 List of Tables .......................................................................................................................... 7 Acronyms ................................................................................................................................ 8 1 Background..................................................................................................................... 9

1.1 Vector Related Diseases of Human Health Importance .......................................... 10 1.1.1 Arboviruses ....................................................................................................... 10

1.1.1.1 Yellow Fever ......................................................................................... 11 1.1.1.2 Dengue fever ........................................................................................ 13 1.1.1.3 Other arboviral diseases transmitted by mosquitoes ............................ 14

1.1.2 Lymphatic Filariasis .......................................................................................... 15 1.1.3 Malaria .............................................................................................................. 16 1.1.4 Tsetse-borne Diseases ..................................................................................... 19 1.1.5 Black Fly-borne diseases ................................................................................. 20 1.1.6 Tick-borne diseases ......................................................................................... 20

1.2 Geography and Ecology of the Area in Relation to Risk from Vector-borne diseases22 1.2.1 Ecology Specific to the Area around Maférinyah and Yindi-Famoriah ............. 24 1.2.2 Ecology Specific to Ile Kaback ......................................................................... 26

2 Scope of the Entomological Study ............................................................................. 28 2.1 Specific Objectives ................................................................................................... 29 2.2 Specific Deliverables ................................................................................................ 29

3 Methodology and Field Activities ............................................................................... 30 3.1 Field Techniques ...................................................................................................... 30

3.1.1 Larval Collection and Rearing .......................................................................... 30 3.1.1.1 Survey Activities for larvae in Maférinyah and Yindi-Famoriah ............ 32 3.1.1.2 Survey Activities for larvae on Ile Kaback ............................................. 35

3.1.2 Larval rearing .................................................................................................... 36 3.1.2.1 Insectary Forécariah ............................................................................. 37 3.1.2.2 Insectary Ile Kaback ............................................................................. 37

3.1.3 Resistance testing ............................................................................................ 38 3.1.4 Human Landing Catches .................................................................................. 41 3.1.5 House Resting Catches .................................................................................... 41 3.1.6 Preservation of Mosquitoes .............................................................................. 42

3.2 Laboratory techniques ............................................................................................. 42 3.2.1 Species Identification ........................................................................................ 43 3.2.2 Detection of knockdown resistance gene ......................................................... 43 3.2.3 Detection of Sporozoites of Plasmodium falciparum ........................................ 44

4 Results........................................................................................................................... 45 4.1 Morphological Identification of Mosquitoes Captured in Field .................................. 45

4.1.1 Human Landing Catches .................................................................................. 45 4.1.2 House Resting Catches .................................................................................... 46

4.2 Morphological Identification of Larvae or Adult Mosquitoes Reared from Larvae .... 46 4.3 Resistance Tests ..................................................................................................... 47 4.4 Laboratory Assays ................................................................................................... 48

5 Summary Discussion on Key Findings and Recommendations ............................. 49



5.1 Malaria and Associated Vectors .............................................................................. 49 5.1.1 Ecology and Potential Vectors .......................................................................... 49 5.1.2 Species Composition and Behaviour ................................................................ 50 5.1.3 Resistance Profile ............................................................................................. 51

5.2 Other Vectors of Medical Importance ...................................................................... 52 5.2.1 Ecology and Potential Vectors .......................................................................... 52 5.2.2 Species Composition and Behaviour ................................................................ 52

6 References .................................................................................................................... 54 7 Appendices ................................................................................................................... 56

7.1 Appendix 1: Standard Operating Procedure (SOP), WHO 1998 – Resistance/Susceptibility Test for Anopheles mosquitoes ................................................. 56



List of Figures Figure 1: Water pot on Ile Kaback .......................................................................................... 11 Figure 2: Duration of Malaria Transmission Season in Guinea .............................................. 23 Figure 3: Rice field in Maférinyah ........................................................................................... 25 Figure 4: Irrigated chilli farm in Yindi-Famoriah ...................................................................... 25 Figure 5: Rice field on Ile Kaback ........................................................................................... 26 Figure 6: Water in drainage channels with rice fields and grasslands ................................... 27 Figure 7: Wetland on Ile Kaback ............................................................................................ 27 Figure 8: Larvae sampling ...................................................................................................... 31 Figure 9: Plastic scoops containing mosquito larvae ............................................................. 31 Figure 10: Sampling larvae in rainwater in Gale .................................................................... 34 Figure 11: Potential breeding sites ......................................................................................... 34 Figure 12: Rice field near Tonguiron ...................................................................................... 36 Figure 13: Insectary Forécariah ............................................................................................. 37 Figure 14: Insectary on Ile Kaback ......................................................................................... 38 Figure 15: Resistance testing in progress .............................................................................. 40



List of Tables Table 1: Yellow Fever Outbreaks in Guinea ........................................................................... 12 Table 2: Behaviours and breeding patterns of most important malaria transmitting vectors in the study area ......................................................................................................................... 18 Table 3: Breeding sites at Maférinyah sub- prefecture ........................................................... 33 Table 4: Breeding sites on Ile Kaback .................................................................................... 35 Table 5: Class of insecticides for mosquito susceptibility ....................................................... 40 Table 6: Mosquito species found in the field .......................................................................... 45 Table 7: Larvae species found in the field .............................................................................. 46 Table 8: Susceptibility/Resistance levels according to WHO ................................................. 47 Table 9: Susceptibility of An. gambiae s.l. .............................................................................. 47



Acronyms An. Anopheles APOC African Programme for Onchocerciasis Control CATT Card Agglutination Test for Trypanosomiasis CCHF Crimean-Congo Haemorrhagic Fever DDT Dichlorodiphenyltrichloroethane HAT Human African Trypanosomiasis HLC Human Landing Catches HRS House Resting Catches IRS Indoor Residual Spraying kdr Knock-down Resistance LF Lymphatic Filariasis LLIN Long-lasting Insecticide Treated Nets LSTM Liverpool School of Tropical Medicine MoH Ministry of Health NMCP National Malaria Control Programme OCP Onchocerciasis Control Programme P Plasmodium PCR Polymerase Chain Reaction RVF Rift Valley Fever SERO Serological Suspects SOP Standard Operating Procedure TBRF Tick-borne Relapsing Fever TL Trypanolysis USSR Union of Soviet Socialist Republics WHO World Health Organisation WNV West Nile Virus YF Yellow Fever



1 Background Vector borne diseases transmitted by arthropods pose an enormous threat to the health of

the population of the Republic of Guinea.

Malaria is the most important of these vector related diseases and is cited as the most

important public health issue, both nationally, and at the local level around the proposed

Project development. The risk from malaria has been extensively discussed in the health

impact assessment and baseline health survey which was completed for the Project [1, 2].

The parasitological study conducted in parallel to this entomological study showed malaria

prevalence in children 6-59 months averaging 32.7% (n=376) in the Forécariah1 prefecture,

with a range in different localities of 12.2-67.4% [1].

Guinea is endemic for Human African Trypanosomiasis (HAT) and bancroftian filariasis.

Onchocerciasis was declared as eradicated in Guinea by the WHO in 2002, but not in

neighbouring Sierra Leone and the possibility of re-introduction cannot be ignored.

Many debilitating and some lethal arboviruses are endemic in Guinea with outbreaks of

Yellow fever occurring sporadically, especially in the southern and eastern sections of the

country that borders on Sierra Leone, Liberia and Ivory Coast. Chikungunya and dengue

fever are also likely to be endemic, but these conditions are generally not recognised as

diagnostic capabilities and the weak health system in the country limits this. It is likely that

many cases are ascribed to malaria when they may be in fact an arboviral disease. This will

especially be true in the more rural areas.

Other arboviruses that may be present include Rift Valley Fever (RVF), West Nile Virus

(WNV), Tahyna and tick borne relapsing fever. These are however likely to be rare.

1 The names of some of the villages mentioned in this report may differ from the official name list established by the project.



1.1 Vector Related Diseases of Human Health Importance Presently, vector related diseases transmitted by mosquitoes cause the majority of morbidity

and mortality associated with communicable diseases in Guinea.

1.1.1 Arboviruses An arbovirus is a virus transmitted between hosts by a vector which may be an insect such

as a mosquito, or another arthropod, such as a tick. As most arboviruses require specialised

assays to detect them, these diseases are not recognised where these facilities are absent.

In a study completed in Guinea, it was reported that 63% of patients with undiagnosed febrile

illness tested positive for arboviruses, which suggests that the burden of disease caused

these conditions is considerably underestimated [3].

During the period from 1978-1991, when the Soviet Union (USSR) and Guinea collaborated

on a Virological and Microbiological Laboratory in Kindia, many strains of arbovirus were

isolated in humans. These included Chikungunya, Dengue serotype 2, Rift Valley fever, and

Crimean Congo haemorrhagic fever amongst others. Dengue 2 viruses were isolated from

mosquitoes.

A recent study to investigate the cause of undiagnosed febrile illnesses in Guinea was

carried out in two hospitals. Diagnostic assays of febrile patients revealed that 63% (30/47)

of patients were infected with arboviruses, including 11 with West Nile, 2 YF, 1 dengue, 8

Chikungunya, and 5 Tahyna infections. Except for YF, these were the first reported cases of

human disease from these viruses in Guinea and the first reported cases of symptomatic

Tahyna infection in Africa. These results strongly suggest that arboviruses circulate and are

common causes of disease in Guinea. Improving surveillance and laboratory capacity for

arboviruses diagnosis will be integral to understanding the burden posed by these agents in

the region [3].

These viruses are generally all transmitted by Aedes group (generally Aedes aegypti or

Aedes albopictus) of mosquitoes. These vectors can sometimes transmit several

arboviruses, often causing confusing mixed epidemics which can be challenging to recognise

and control effectively.

The one exception is the West Nile virus which is transmitted by mosquitoes from the Culex

group of mosquitoes.



1.1.1.1 Yellow Fever Yellow fever (YF) is transmitted by species of Aedes mosquitoes, mainly Aedes aegypti and

to a lesser extent Aedes albopictus. Aedes spp. are primarily forest mosquitoes, however

Aedes aegypti and Aedes albopictus live in close association with man, in any kind of human

settlement. Both, Aedes species breed in any small water collection including tree holes,

banana trees and plant axils. Rapid urbanization, the accumulation of water-retaining waste

products such as plastic containers and tin cans, coconut shells and increased domestic

water storage, for example in large open drums, cement tanks and earthenware vessels,

have all increased the availability of Aedes breeding sites.

Ile Kaback and the broad study area represent ideal breeding grounds for this group of

mosquitoes. Figure 1 depicts a ceramic water storage pot which is commonly used to store

drinking water. These are ideal breeding grounds for Aedes mosquitoes.

. Figure 1: Water pot on Ile Kaback

Aedes aegypti and Aedes albopictus bite during the day outside often in shady areas,

although Aedes albopictus will enter houses and bite during the night. The mostly exophagic

(outdoor feeding) and exophilic (prefers to rest or stay outdoors) behaviour means these



vectors are not susceptible to control by insecticide treated bednets (ITNs) or indoor residual

spraying (IRS).

YF is the most commonly reported disease transmitted through these mosquitoes.

Table 1 outlines recent YF outbreaks in Guinea [4]. It is noteworthy that the only outbreak in

Maritime Guinea occurred in Boké.

Table 1: Yellow Fever Outbreaks in Guinea

Year/Month of outbreak Location of outbreak Suspected cases;

confirmed cases Deaths

2000 15 districts in north-western Guinea

512 suspected cases; confirmed cases in 10 districts

190

2001 Conakry 18 suspected cases 2 confirmed cases

2

2001 N’Zérékoré 11 suspected cases - 2003, January Macenta and Kérouané

prefectures 43 suspected and confirmed cases

24

2004, December Faranah 6 suspectec cases 6 confirmed cases

-

2004, December Mamou 4 confirmed cases 3 2004, December Dalaba 3 confirmed cases 1 2005, November Boké, Kankan,

N’zérékoré, Faranah and Conakry

9 suspected cases -

2005, December Nationwide 114 suspected cases 23 confirmed cases

26

2008, August and September

N’zérékoré 14 suspected cases 1

2008, December Faranah 21 suspected cases 2 confirmed cases

3

2009, August Faranah 21 suspected cases 3 2010, January Mandiana 1 confirmed case - 2010 Faralako 6 suspected cases -

In response to YF outbreaks, several mass vaccination campaigns have been performed in

affected areas. In 2000, a mass vaccination campaign targeted 15 districts in Guinea. In

2003, ~600,000 people were vaccinated in Macenta and nearby prefectures. As a response

to the 2005 outbreak in Boké, a mass vaccination campaign was initiated in 4 high risk

districts (Boké, Boffa, Gaoual and Koundara). A campaign was also carried out in N’zérékoré

with reported 92.5% coverage except in Bounama sub-prefecture, Urbain Commune and the

Ivorian Refugee Camp of Kouankan 2 where a low coverage rate was reported (45%). In

December 2008, on the basis of an outbreak investigation, a reactive mass vaccination

campaign, planned targeting 60,485 people, was carried out in some strategic areas in the



health district of Faranah. In 2010, the MoH has decided to vaccinate the entire population of

the Mandiana prefecture. There was no evidence that Forécariah prefecture had benefitted

from any past vaccination campaigns.

The risk of introduction of arboviral diseases in the proposed port area as a direct impact

from the Project is discussed extensively in the health impact assessment (HIA). The port

activity has the potential to introduce diseases from areas outside of the country and the rail

route has the potential to transfer the disease from the interior of the country to the coastal

region. There may also be an increased movement of people into the study area with

potential for introduction of the virus into an area where the environment is conducive for the

proliferation of Aedes aegypti. As described later in the report the vectors are present in the

study area and the environment conducive to support the breeding and proliferation.

1.1.1.2 Dengue fever Dengue, like Yellow fever, is transmitted by mosquitoes of the genus Aedes (mainly Aedes

aegypti and to a lesser extent Aedes albopictus). There are four distinct, but closely related,

serotypes of the virus that cause dengue (DEN-1, DEN-2, DEN-3 and DEN-4). Dengue is

regarded as one of the most important arboviral diseases globally whose range has

extended significantly in the past few years [20].

The virus is transmitted to humans through the bites of infected female mosquitoes. After

virus incubation for 4–10 days, an infected mosquito is capable of transmitting the virus for

the rest of its life. The virus can also be transmitted via the eggs of an infected mosquito

resulting in infected mosquitoes when they emerge. These eggs can also desiccate for a

period time and emerge under the ideal environmental conditions.

Infected humans are the main carriers and multipliers of the virus, serving as a source of the

virus for uninfected mosquitoes. Patients who are already infected with the dengue virus can

transmit the infection for 4–5 days; maximum 12 days via Aedes mosquitoes after their first

symptoms appear. Female Aedes aegypti bite multiple people during each feeding period.

Increased international travel and especially across shipping routes has led to the mixing of

different strains of the disease, causing more severe clinical symptoms in individuals

exposed to more than one strain [5].



There have been recent epidemics in countries neighbouring Guinea, the most recent being

in Senegal and since the vector is present there is a likelihood of dengue becoming more

prevalent in Guinea with the possibility of epidemics occurring. This is described in more

detail in the HIA.

The fact that once a mosquito is infected it can transmit dengue virus for life, coupled with

the knowledge that Forecariah is suitable for Aedes breeding is cause for concern especially

considering the high volume of movement of people from differing geographical locations in

and out of the area related to the mining project.

1.1.1.3 Other arboviral diseases transmitted by mosquitoes Other arboviruses which are medically important in Guinea are Chikungunya and West Nile

Virus (WNV).

Chikungunya is a viral disease (genus Alphavirus) which is transmitted to humans by

infected mosquitoes of the genus Aedes– including Aedes aegypti and Aedes albopictus.

Chikungunya is rarely fatal. Symptoms are acute and characterised by fever, rash, and

incapacitating bone pain. It may be confused clinically with dengue although the bonce ache

is unique to Chikungunya. Symptoms are generally self-limiting and last for 2–3 days. The

virus remains in the human system for 5-7 days and mosquitoes feeding on an infected

person during this period can also become infected [6].

Humans serve as the Chikungunya virus reservoir during epidemic periods. Outside these

periods the main reservoirs are monkeys, rodents, birds, and other unidentified vertebrates.

In Guinea, Chikungunya is endemic and there is likely to be high-level and continuous

transmission to largely immune populations, and small rural outbreaks or sporadic cases.

Studies in Guinea (albeit with a small sample size) have reported that Chikungunya may

account for as many as 17% of undiagnosed fevers [3].

Due to the likelihood of increased suitable breeding sites for Aedes because of the Project’s

activities, there is a strong possibility of Chikungunya increasing in prevalence at the local

level. Many expatriate personnel may not have been previously exposed to Chikungunya,

and the potentially naïve local population can lead to a localised epidemic in the area which

has the potential to extend past the immediate area. While the disease is generally not life



threatening, its debilitating nature may increase morbidity in the area and cause absenteeism

amongst employees. There is also the potential for reputational consequences to the

company.

West Nile virus (WNV) is an avian virus that can cause fatal disease in some species of

mammals, reptiles and birds. Most clinical cases occur in humans and horses. Approximately

80% of infected humans remain asymptomatic; 20% have flulike symptoms. Less than 1%

develop meningitis, encephalitis or acute paralysis, with some of these cases leading to

death or resulting in permanent impairment.

WNV is primarily transmitted by mosquitoes, and a large number of different species are

efficient vectors. Culex quinquefasciatus, endemic in Guinea, and found breeding in many

types of sites, is an efficient vector, as is Aedes albopictus. Recent studies have reported

that WNV may account for as much as 23% of undiagnosed fevers in Guinea.

Culex spp. can breed in a wide variety of habitats, but some show a preference for foul or

heavily polluted water which is often associated with peri-urban or urban locations. They can

have a specific preference for pit latrines. They are thus commonly found in areas of polluted

water sources and were expected to occur commonly in the towns of Maférinyah and

Forécariah.

1.1.2 Lymphatic Filariasis

Guinea has been declared free of Onchocerciasis and Dracunculiasis but lymphatic filariasis

(LF) is still prevalent in Guinea. This disease is caused the parasite Wuchereria bancrofti and

is transmitted by Anopheles species principally Anopheles gambiae ss, Anopheles arabiensis

and Anopheles funestus.

The community directed Ivermectin® mass-treatment campaigns supported through the

African Onchocerciasis Control Programme has led to very low levels of disease

transmission. While the drug does not kill the adult worms, this together with the introduction

of ITNs for control of malaria, have brought about decreases in rates and intensity of

microfilaraemia and this lowered transmission risk.



In studies on molecular forms of Anopheles gambiae ss, An. gambiae M was significantly

correlated with LF, and 2.5 to 3 times more prevalent in the high LF zone than low to medium

zones. Thus areas with predominantly M form should be regularly assessed as this disease

could be eradicated from Guinea [7].

1.1.3 Malaria Malaria in humans is caused by four main species of the protozoa Plasmodium (P.); P.

falciparum, P. ovale, P. vivax and P. malariae. In Guinea, 98% of infections are attributed to

P. falciparum, the most dangerous form, which can be fatal within a week in people with

inadequate immunity, especially children, pregnant mothers and those from non-endemic

areas. The disease is transmitted from human to human by female Anopheles mosquitoes.

Although malaria is endemic throughout Guinea, no reliable regional or national estimates of

parasitaemia exist, nor are entomologic data related to malaria transmission widely available.

The NMCP (and the Global Fund) has characterized regions in Guinea as being

hyperendemic, holoendemic, mesoendemic and hypoendemic, although recent studies or

data on which these characterizations are based are lacking.

National statistics in Guinea show that among children less than five years of age, malaria

accounts for 31% of consultations, 25% of hospitalizations, and 14% of hospital deaths in

public facilities. This estimate does not include malaria cases seen in the community or in

private facilities. In addition, most malaria cases reported in national statistics are clinically

diagnosed, and therefore may not accurately reflect the true malaria burden. A recent pilot

study in two regions of Gambia found that of 429 clinically diagnosed cases of malaria in

hospitals, health centres, and health posts, only 26% actually had malaria according to

microscopy (unpublished data, National Public Health Institute and Medical Research

Council in the Gambia). This is considered relevant as it is in the West African Region.

According to national health statistics, the incidence morbidity rate for malaria in Guinea is

148/1,000 population.

Guinea has year-round malaria transmission with peak transmission from July through

October in most areas (NMCP Strategy 2006–2010).

The major vectors in the country are members of the Anopheles gambiae complex including

Anopheles gambiae ss and Anopheles arabiensis. Anopheles gambiae s.s. is highly



anthropophilic (prefers to feed on humans). Anopheles funestus is also an important vector

and exploits permanent or semi-permanent breeding sites such as marshes or rice fields. Its

population density peaks in the dry season, extending malaria transmission by relay after An.

gambiae and An. Arabiensis populations have declined due to lack of breeding sites

favourable to them. These behaviours are discussed in more detail in Table 2.



Table 2: Behaviours and breeding patterns of most important malaria transmitting vectors in the study area

Anopheles species

Vector Importance

Resting location and distribution Feeding pattern Host preference Preferred breeding sites General Control

Recommendations An. gambiae Most important

competent vector in sub-Saharan Africa

Tend to rest indoors immediately after feeding followed by movement to outdoor resting sites after 12-24 hours. Prefers areas of higher relative humidity than An. arabiensis.

Mainly late indoors with higher potential for this than An. arabiensis.

Mainly humans Most often in fresh water ephemeral habitats in shallow, open and sun-lit ponds, puddles, hoof prints and other ground depressions devoid of marginal vegetation. Water is characteristically turbid, low to moderate organic content, with moderate algal growth. Rice fields are often ideal.

Integrated vector control with a focus on source reduction, ITNs and IRS.

An. arabiensis Competent vector Indoors and outdoors. An. arabiensis appears more commonly in arid, sparsely vegetated savannahs. Prefers areas of lower humidity.

Mainly late indoors and outdoors

Humans and animals Temporary pools. Rice fields. Integrated vector control with a focus on source reduction, ITNs and IRS are important but less effective. Larvaciding is more important. This species is considered more difficult to control due to its resilience in harsher environments and adult feeding characteristics.

An. funestus Competent vector with extremely important role extending disease transmission periods. a generally robust and long-lived species

Mainly indoors. Long flight range Mainly late indoors Mainly humans Semi-permanent and permanent clear water, especially with emergent vegetation. Common in swamps, slow streams (particularly water under shade), edges of ditches and the grassy edges of slow moving rivers.

Integrated vector control with a focus on source reduction, ITNs and IRS. Larvaciding is more important. This species is important as it can emerge as the main disease vectors as An. gambiae effects wane- thus have a relay effect and sustaining transmission due to different vector behaviours.

An. melas Important vector in area with mangrove swamps. Can also extend transmission season. Not a highly effective vector but role in study area will be important to monitor.

Mainly outdoors Mainly late outdoors Humans and animals Coastal area, brackish water, mangroves and salt marsh.

Integrated vector control with a focus on source reduction. ITN and IRS less effective. Larvaciding can be useful.



A recent study of malaria vectors in south-eastern Guinea found Anopheles gambiae s.l.

(77%) was the most common Anopheles collected, followed by An. funestus (20%).

Household surveys showed different parasite rates in the three areas studied: 78.2% of

children had parasitaemia in Sombili, 45.8% in Timbi Madina and 16.7% in the urban sector

of Labé. The recent parasitological study conducted for Rio Tinto as part of the HIA also

showed considerable variation in parasitaemia between study sites independent of

topographical features such as elevation above sea level. Beyla and Macenta prefectures

showed a range in different sentinel sites as measured in children under 5 years of 48.2-

85.9% and Forécariah showed a range between sites of 12.2-67.4% [1].

Recognition of local variations in the malaria epidemiology is critical since identification of the

factors responsible for a low prevalence in one village, but a high one in a neighbouring

community may indicate a possible control measure. Local variations in the epidemiology of

malaria must also be taken into account when any kind of malaria intervention is instigated.

1.1.4 Tsetse-borne Diseases The only disease known to be transmitted to humans by tsetse flie,s Human African

Trypanosomiasis (HAT), also known as sleeping sickness, is a vector-borne parasitic

disease. The parasites concerned are protozoa belonging to the Trypanosoma genus. They

are transmitted to humans by tsetse fly (Glossina genus) bites after the tsetse have acquired

their infection from human beings or from animals harbouring the human pathogenic

parasites. In Guinea, the human infective subspecies of Trypanosoma brucei (T.b) causing

HAT is T.b. gambiense and the disease is endemic especially in the coastal region with local

transmission in Forécariah prefecture. These trends have been described in detail in the HIA

[2].

Recent research on HAT shows that the fauna of tsetse flies in Guinea includes at least 8

species, two of which are vectors of HAT. These include G.(N.) palpalis and G.(N.)

tachinoides, of which the latter is the vector of animal trypanosomiasis ("nagana" cattle

disease) as well [8].

In a study conducted in the Forécariah mangrove focus in Guinea, patients with confirmed

HAT and serological suspects (SERO) were identified through mass screening of the

population using the card agglutination test for trypanosomiasis (CATT). Analysis of the

samples collected during the follow-up of HAT patients and SERO was performed with



polymerase chain reaction (PCR) and trypanolysis (TL) serological tests in order to clarify the

role played by these individuals in the epidemiology of HAT. PCR positivity was higher in TL⁺

than in SERO TL⁻ (50% vs. 18%, respectively). Whereas CATT plasma titres decreased both

in treated HAT patients and SERO TL⁻, SERO TL⁺ maintained high CATT titres. 4 out of 17

SERO TL⁺ developed HAT during the study. These results strongly suggest that SERO TL⁺

individuals are asymptomatic carriers. In the context where disease prevalence is sufficiently

low, treating SERO TL⁺ individuals may thus be of crucial importance in order to interrupt

transmission [9].

1.1.5 Black Fly-borne diseases The only human disease known to be transmitted by blackflies is Onchocerciasis.

Onchocerciasis is a parasitic disease caused by the filarial worm Onchocerca volvulus.

Following larvicide applications commencing in West Africa in 1974 coupled with annual

large-scale community directed ivermectin® distribution since 1987, blackfly infectivity was

greatly reduced and in 2002 onchocerciasis was officially (according to WHO) eradicated in

Guinea [10].

Although onchocerciasis has been eradicated the Simulium vectors are not. The most widely

distributed species of Simulium in Guinea were S. sirbanum and S. damnosum s.s.,

associated with savannah areas, recorded from all river basins S. soubrense 'Chutes Milo'

form in Guinea, both 'Konkouré' and 'Menankaya' forms of S. konkourense occurred

predominantly in Guinea [11].

Prior to eradication, onchocerciasis was mostly prevalent in north and northwest Guinea

close to large rivers, but away from Maritime Guinea. However, the disease still occurs in

Sierra Leone, and although the rates have drastically reduced in the past few years, the

disease still occurs along the coast. This is important for the study area as there is reported

to be significant movement of people between the two countries and the proposed Project

may serve as an attractor with increased movements and disease introduction.

1.1.6 Tick-borne diseases The epidemiology of tick borne diseases in Guinea is not well studied. However tick borne

diseases caused by arboviruses which are likely to cause considerable morbidity and



mortality are Crimean-Congo haemorrhagic fever (CCHF) and tick-borne relapsing fever

(TBRF).

Virus-tick-vertebrate host relationships are highly specific and less than 10% of all tick

species (Argasidae and Ixodidae) are known to play a role as vectors of arboviruses.

However, a few tick species (e.g. Ixodes ricinus, Amblyomma variegatum) transmit several or

many (I. uriae) tick-borne viruses.

Crimean-Congo haemorrhagic fever (CCHF) has a widespread distribution, closely

approximating the known global distribution of the Hyalomma spp ticks which transmit the

disease [12]. The disease is caused by a Nairovirus, and is a zoonosis with infected animals

remaining asymptomatic but with a high case fatality rate in humans [13].

Tick-borne relapsing fever (TBRF) is caused by several species of Borrelia spirochetes

and is transmitted to humans through the bites of soft ticks of the genus Ornithodoros. Wild

rodents and insectivores are common reservoir hosts. The extent of relapsing fever caused

by infection with B. crocidurae, transmitted by O. sonrai ticks, and its effects on public health,

has only just begun to emerge. Although there is no Guinea specific data, research in

neighbouring countries such as Senegal, Mali, Mauritania, and the Gambia where this tick is

endemic, showed 2%–70% of animal burrows to be inhabited by this tick vector, and an

average of 31% of ticks are infected by B. crocidurae [14]. TBRF is underdiagnosed in most

disease-endemic areas, where blood smears are screened only for malaria parasites. B.

crocidurae causes high fever, frequent neurologic complications, and up to 9 recurrences

over several months.

African tick bite fever is an acute febrile illness that is frequently accompanied by

headache, prominent neck muscle pain, inoculation sores (eschars) and regional lymph node

swelling. The disease is caused by Rickettsia africae which belongs to the spotted fever

group rickettsia and is transmitted by ungulate ticks of the Amblyomma genus in rural sub-

Saharan Africa [15]. Whereas reports on African tick bite fever in indigenous populations are

scarce, the number of reported cases in travellers from Europe and elsewhere has recently

increased significantly. African tick-bite fever is an important emerging infectious disease



1.2 Geography and Ecology of the Area in Relation to Risk from

Vector-borne diseases Forécariah prefecture covers an area of 4200 km2 situated in south-east Guinea. The

population is estimated at around 200,000, most of whose livelihood is supported by

subsistence farming or fishing. The region is characterised by many rice growing areas and

irrigated vegetable and chilli farming.

The prefecture of Forécariah is a coastal lowland zone bordering the Atlantic Ocean and

forming part of Maritime Guinea, which is defined as one of the four ecologically distinct

areas of Guinea. The land comprises more or less sandy, marshy alluvial with estuaries and

deltas, colonised by mangrove vegetation. The plains characterising this zone possess vast

cultivated estuarine rice fields (Bogoni) protected by a dense row of mangroves. Thousands

of farmers make these plains the rice granary of Guinea. In addition, there is extensive

irrigated farming of chillies in the area around Maférinyah and Yindi-Famoriah.

The coastal area is the wettest part of the country with annual precipitation exceeding

4,000mm. The rainy season in this part of the country lasts for seven to eight months (from

March, April, through October to November). The average temperature during the hottest

period of the year (March/April) is 27-30oC, however temperatures reach 36oC. The lowest

daytime temperatures are around 23oC with temperatures falling to 21oC at night. The area is

humid. The climatic features allow for year round transmission of malaria but the peak

malaria transmission is present for 8-9 months of the year, as shown in Figure 2 [21].



Figure 2: Duration of Malaria Transmission Season in Guinea

All of these factors including the associated habitat modification from farming lead to

conditions favouring breeding sites of Anopheles mosquitoes. Cattle and domestic animals

are not common in the study area.

In peri-urban areas in the study area (Maférinyah and Forécariah) poverty associated with

rapid population growth has led to concentrations of people without the necessary

infrastructure for the safe storage and distribution of water and drainage of wastewater.

Households store water in used containers and with no system for rubbish disposal old

containers, tins, tyres etc. provide ample breeding sites for mosquito vectors especially

Aedes spp. The poor sanitation in the area may also promote the proliferation of Culex

species such as Cx. quinquefasciatus.



1.2.1 Ecology Specific to the Area around Maférinyah and Yindi-Famoriah

There is an abundance of rice fields in these locations. Those lying close to the ocean

contain brackish water, which would be suitable for An. melas, however those further inland

contain freshwater and are suitable for other members of the Anopheles gambiae complex

as well as Anopheles funestus.

Many of the rice fields are irrigated with water from small creeks, but the water does often not

flow in the irrigation ditches providing many potential breeding sites for An. funestus (Figure

3). However, in some cases the water was very stagnant and only suitable for Culicines to

breed in. On the chilli farms, the irrigation areas has mainly stagnant water which were also

more suited to Culicines (Figure 4). Marshy areas and ponds with grassy edges are

numerous in the environment, again providing potential for Anopheles gambiae and

Anopheles funestus to breed.

Maférinyah and Forécariah are urban centers and are reported to be growing rapidly. The

increased urban movement with no corresponding increase in sanitation or water supply will

likely provide ample breeding sites for Aedes aegypti and Culicine vectors.

The mangrove swamps found along much of the coast especially in the river mouths, where

the water is brackish (part river water, part see water) are habitats of Glossina palpalis, which

is likely to the most important vector for the transmission of HAT in the study area. Fresh

water swamp forest found in the areas surrounding the rice fields is also favoured by G.

palpalis. Although much of the land has been altered for agriculture there are still

considerable areas of savannah further inland with trees forming open woodland with tall

grass. Glossina longipalpis occurs in the southern parts of this zone. Further north, G.

morsitans replaces it. Rivers and streams along which evergreen trees grow form the habitat

of G. palpalis and G. tachinoides[16]. The fact that diagnosis of HAT is difficult and the

majority of the population live a long way from health facilities means that considerable under

diagnosis is likely to occur and the disease may be a far greater problem than realised.

The rural population of Guinea may be at risk for tick-borne rickettsioses but this may be an

unrecognised cause of fever.



Figure 3: Rice field in Maférinyah

Figure 4: Irrigated chilli farm in Yindi-Famoriah



1.2.2 Ecology Specific to Ile Kaback Ile Kaback is situated in low lying areas enveloped with mangrove thickets. Settlements are

present along a small network of roads and represent small hamlets in the main. The rest of

the land around the human habitation is almost entirely developed into rice fields. Many of

the rice fields are permanently wet, being fed by rainwater, underground springs and small

creeks. There is no flow of water in the rice fields and they thus provide ideal breeding sites

for many mosquito species, including the main malaria vector species, An. gambiae.

In contrast to Forécariah, the amount of discarded rubbish was not found to be as significant

a problem in the villages surveyed for breeding sites for Aedes aegypti. However, water is

stored in receptacles that could serve as effective breeding sites as shown in Figure 1.

However, this situation could change with an influx of people from outside or with

commencement of construction which often leads to discarded drums, tyres and other

suitable breeding sites being formed.

Figure 5: Rice field on Ile Kaback

The rice fields in Ile Kaback are generally surrounded by small drainage channels and where

there is no active rice cultivation wet grasslands predominant. These are ideal breeding

grounds for An. funestus as shown in Figure 6.



Figure 6: Water in drainage channels with rice fields and grasslands

Figure 7 depicts a wetland on Ile Kaback which may also be suitable habitat for breeding of

An. funestus.

Figure 7: Wetland on Ile Kaback



2 Scope of the Entomological Study The scoping study HIA conducted in September 2010 reported that there was limited data on

the local entomology in the area. This data gap was considered important due to the

proposed port development and how any demographic or ecological change could impact on

the transmission of vector related diseases.

Thus the entimology study had the had the broad objective of understanding; i) what

common disease transmitting vectors were present in the area; ii) how the proposed Project

may change the risk of disease transmission based on changes to the local ecology; and iii)

the insecticide sensitivity pattern of locally important malaria vectors.

It was the intent to then use this information to support the HIA and subsequent development

of evidence based mitigation and management measures to avoid or limit these impacts. It

was anticipated that the Project development may cause changes to the environment,

ecology and demographics in the area and thus influence local patterns of vector related

disease transmission. Further, it was the intent that the study would support a component of

the evidence to support the design of local vector control strategy for workplace health

management as well as community health considerations.

While the current assessment is limited to a specific time period of the year and will not

provide a detailed analysis of the local disease transmission and vector patterns, it does

provide a snap-shot of the local ecology and entomology. This allows the qualitative

determination of the likely patterns of disease transmission in the area and adequate

evidence to base at least initial vector control activities, and support decision making related

to design elements of the Project. Ideally, when the Project progresses into pre-development

an entomological surveillance programme should be established to determine vector

abundance and behaviours at different seasons and linked to changes in the environment.

This should include specific surveillance in the wet and dry season as well as methods

applied to monitor endophilic (indoor biting), exophilic (outdoor biting) and day biting and

night biting mosquitoes and other vectors such as tsetse flies, blackfly and ticks.



2.1 Specific Objectives Due to the time periods available for the entomology survey it was important that the

objectives were focussed to a specific outcome.

The primary objectives of the study included:

• To determine the predominant mosquito species which are present, and which could

potentially occur, in the study area in Forécariah prefecture. It was therefore decided

to concentrate efforts on sampling mosquito larval populations from a variety of

aquatic habitats, as well as using a range of mosquito adult sampling methods to

determine what vectors of different diseases existed in the area.

• To determine the susceptibility of malaria mosquito vectors found in the study area to

different classes of insecticides. The resistance study was conducted using a

representative insecticide from three major groups of insecticides, namely

pyrethroids, carbamates and organophosphates. The study was completed on first

generation mosquitoes from larvae collected from different breeding sites in the area

to ensure that the age of the mosquitoes was known.

Secondary objectives included:

• To determine the main species of mosquitoes which are responsible for malaria

transmission in the area.

• To establish what potential mosquito vectors of other diseases occur in the area, for

example Aedes and Culex.

• To collect sufficient information on resistance of mosquito disease vectors to be able

to produce an evidence-based control strategy.

• To verify the mechanism of resistance if any is found.

• To confirm which molecular forms of the malaria vector Anopheles gambiae, if found,

are present.

2.2 Specific Deliverables The following were deliverables from the assessment:

• To describe the local entomology of important to human health risk

• To support evidence for the HIA to define evidenced based management and

mitigation measures.



3 Methodology and Field Activities 3.1 Field Techniques The following mosquito collection methods were used in support of the assessment:

• Larval collection and rearing;

• Human landing catches (HLC); and

• House resting catches (HRC)

These are described in more details below.

3.1.1 Larval Collection and Rearing All potential breeding sites within a 3.5km radius of sentinel sites used in the Port baseline

health survey on Ile Kaback were sampled (Figure 8) [1]. The distance was limited by the

size of the island. For the mainland, breeding sites within a radius of 10kms of each sentinel

site were sampled.

Mosquito larvae of any species were collected using plastic scoops and were individually

pipetted from the scoop to separate each larval species morphologically (as some species

are predatory on others). Collections from each breeding site were kept separate and

labelled at source (Figure 9).

With initial surveys in large potential breeding sites such as rice fields sampling by dipping

was carried out every five metres around the perimeter and along several transects within

the body of the rice field. There is no hard and fast rule to define a potential breeding site as

negative, but in rice fields larvae generally prefer to inhabit the grassy edges and may be

localised to the sides closest to human habitation. However, if the breeding site is significant

it is unlikely to be misclassified as negative for larvae with the above procedure.



Figure 8: Larvae sampling

Figure 9: Plastic scoops containing mosquito larvae



3.1.1.1 Survey Activities for larvae in Maférinyah and Yindi-Famoriah Approach: Although the largest potential breeding sites in these sites were rice fields, there was also

considerable conversion of land to chilli farming which covered areas of up to two hectares

and was always irrigated. Rice fields and irrigation ditches of chilli farms were sampled for

mosquito larvae.

Most of the small villages within the vicinity of the rice fields and chilli farms were kept clean,

however, breeding was found in many vessels left lying around the villages. Mostly these

were unused (i.e. not water storage containers in use) discarded vessels especially pottery

cooking pots, broken plastic receptacles and buckets, old tyres and scrap metal. In one

village, Fanye, one household had a large 400 litre plastic storage box, and another had a

large cement water storage tank fed by rain water wherein mosquito larva were found. Other

cement water storage tanks were found but at that time were dry. A number of wells were

inspected but larvae were absent at the time of the survey. In Fanye, high concentration of

mosquito larvae was found in a duck pond belonging to one residence. Other potential

breeding sites were pits dug outside households and especially brick making areas, where

borrow pits had been dug.

Findings: Breeding sites were only found in the Maférinyah area (Figure 10 and Figure 11). More than

50 potential breeding sites were located in the week in the field at these locations. These

sites varied from chilli farms averaging 1-3 hectares, to rice fields which varied from 1-100

hectares, and smaller communal fields. The description and number of each site type visited

is shown below in Table 3.

No Anopheles larvae were found in any of the breeding sites within the vicinity of Yindi-

Famoriah. This should not be considered as an indication that breeding never takes place in

this area, but rather that during the week of the survey it was not possible to locate localised

breeding sites. It must be recognised that in the days prior to the survey and during the

survey heavy rain was experienced in the evening which may have flushed out these

breeding sites.

In the Maférinyah area several breeding sites were found, although, all except the duck

pond, had very low densities of Anopheles larvae averaging one larva per 20 dips (each dip



is approximately 0.5 litres). The duck pond had an exceptionally high density of Anopheles

larvae averaging nine larvae per dip.

Table 3: Breeding sites at Maférinyah sub- prefecture

Sentinel Site Type of potential breeding site

Number sampled Number found with mosquito

larvae

Mosquito genus found

Maférinyah

Rice field 23 1 Anopheles, Culex Chilli farm 5 1 Anopheles, Culex Duck pond 1 1 Anopheles, Culex Marsh 3 2 Anopheles, Culex Well 2 0 Concrete water storage tank

2 1 Anopheles, Culex

Plastic water storage tank

3 3 Culex, Aedes

Brick making pits 2 1 Anopheles, Culex Discarded vessels 18 12 Culex, Aedes

Yindi-Famoriah

Rice field 11 0 Chilli farm 1 0 Swamp 1 0 Discarded vessels 13 8 Culex, Aedes

Resistance testing for Yindi-Famoriah could not be set up due to the lack of larvae found in

that area. Resistance testing for Maférinyah used adults emerging from mixed samples of

Anopheles gambiae s.l. collected from the duck pond at Fanye together with two marshes

along the road from Maférinyah to Touguiyiré, and another at a chili farm close to

Maférinyah.



Figure 10: Sampling larvae in rainwater in Gale

Figure 11: Potential breeding sites



3.1.1.2 Survey Activities for larvae on Ile Kaback Approach: All large scale potential breeding sites in this area were located in the rice fields (Figure 12).

There was no chilli farming and there were no marshy areas not associated with rice fields.

The population lived in small villages which were mostly kept clean, however each village

was checked for mosquito breeding in any water storage containers and discarded vessels.

Matakang was more prone to having rubbish strewn around, however, at the time of the

study no active breeding sites were found in any of the containers inspected.

Findings: Larvae were only isolated in the rice fields that surrounded the settlements.

Anopheles gambiae s.l. was used for resistance testing along with the few Anopheles

funestus found as larvae.

No larvae of Aedes aegypti were found although one adult was caught biting during the day

and the people living nearby reported the presence of day biting mosquitoes at certain times

of the year.

Table 4: Breeding sites on Ile Kaback

Sentinel Site Type of potential breeding site

Number sampled Number found with mosquito

larvae

Mosquito genus found

Manké Rice field 23 1 Anopheles, Culex Discarded vessels 18 12

Matakang Rice field 11 0 Anopheles, Culex Discarded vessels 13 8



Figure 12: Rice field near Tonguiron

3.1.2 Larval rearing The larvae collected were reared to adults in temporary insectaries. On return from the field,

Anopheline larvae morphologically identified as Anopheles gambiae spp. were separated

from other larvae. However, they were pooled with Anopheles gambiae spp. larvae from

other breeding sites within the sentinel site boundary to ensure a genetic mix was available

for resistance testing (larvae from one site may be the progeny of one or few females

unrepresentative of the general population).

Anopheles funestus larvae from all breeding sites within the sentinel site boundary were

pooled separately. Other Anophelines and Culicine larvae were pooled separately

morphologically for rearing.

All collection information was recorded on the larval rearing bowls. Larvae were fed on

imported Tetramin crushed to a fine powder. Larvae were reared in their original source

water which was collected at the time of larval sampling. If the rearing bowls retained too

much sediment the water was replaced with freshly collected water from the original source.

Larval bowls were kept covered with untreated (insecticide free) mosquito netting to ensure

adult mosquitoes which might enter the rooms did not lay eggs in the bowls. Larvae which

developed into the pupal stage were removed in the morning and evening and placed in

pupal cups into a cage suitable for emerging into adults. Adult mosquitoes were fed on

sucrose solution soaked on cotton wool, and were kept from drying out by partially covering

the cages with damp towels.



3.1.2.1 Insectary Forécariah In Forécariah, the annex to a bedroom was utilised as an insectary (Figure 13). Care was

taken to ensure the table legs were continuously submerged in water to prevent attack of the

caged mosquitoes by ants and the rearing table was kept away from the wall for the same

reason. Once adults had emerged they were kept for 2-5 days and the females were then

used for resistance testing and the males discarded.

All Culex and Anopheles species which were not vectors of any known disease were killed

and preserved for formal laboratory identification.

Figure 13: Insectary Forécariah

3.1.2.2 Insectary Ile Kaback On Ile Kaback, the entomologists’ bedrooms in the guest house was initially used as a

temporary insectary, which due care taken to avoid knock down insecticides and ITNS in the

room (Figure 14). Later when the bedrooms were required by another team the entomology

team hired a bedroom in a local house in the village. For the last stage of the work the room

in the guest house were again used. All rooms proved completely satisfactory, the main

concerns being to ensure protection of the emergent adults from ant attack and to ensure the

rooms remained cool, but relatively humid.



Figure 14: Insectary on Ile Kaback

3.1.3 Resistance testing Resistance testing was carried out according to the procedure detailed in Appendix 1 andd

shown in Figure 15.

The reared mosquitoes of medical importance were tested against three classes of

insecticide as described in



Table 5. These were tested using World Health Organisation (WHO) standardised

insecticide-impregnated papers in specific prepared tubes. WHO provides a choice of

alternative insecticides for each class of insecticide. The insecticide impregnated papers

chosen were for the insecticides most commonly in use for malaria control and felt to be

most relevant to this study.

The mosquitoes were exposed for an hour and then returned to containers containing cotton

wool soaked in sucrose solution and maintained in ant protected surroundings with adequate

humidity for the remaining 24 hours of the test. At the end of each resistance test all

mosquitoes were preserved according to the procedure described below.



Table 5: Class of insecticides for mosquito susceptibility

Insecticide class Representative insecticide used

Concentration Control paper used

Pyrethroid Deltamethrin 0.05% PY-Control (silicone oil)

Carbamate Bendiocarb 0.1% OP/C-Control (olive oil)

Organophosphate Fenitrothion 1.0% OP/C-Control (olive oil)

Figure 15: Resistance testing in progress



3.1.4 Human Landing Catches Human landing catches (HLC) were carried out at the Manké sentinel site area on Ile Kaback

and in the Fanye area close to the Maférinyah sentinel site. They were planned for Matakang

but the area was not accessible at the scheduled time due to heavy rain. Yindi-Famoriah was

excluded based on the absence of larvae and reports from the local population of minimal

mosquito activity. This was confirmed through a short assessment of indoor resting adult

mosquitoes using knock down sprays.

The catches were conducted to sample for mosquitoes that will readily feed on humans. The

HLC were performed indoors and also in the area outside the same house.

The house owners willingly gave their informed consent to this activity. All three

entomologists participated in the catches. Catches commenced at 6pm and continued until

6am. Catches were separated according to hourly interval and outside catches were kept

separate from indoor catches. The catchers rotated at two hourly intervals throughout the

night to reduce effects on the catches caused by individual mosquito attractiveness. It should

be noted that a far more rigorous rotation schedule together with many more nights of

catching would have been necessary to determine biting cycles of the vectors caught. Hourly

catches were maintained to explore any indication of vectors only biting during a specific

period.

3.1.5 House Resting Catches House resting catches (HRC) are used to sample indoor resting (endophilic) mosquitoes.

Mosquitoes which rest indoors are usually also indoor biting (endophagic) but may be

outdoor biting (exophagic). Mosquitoes resting anywhere indoors, usually on walls (except

where indoor residual spraying occurs), under beds, on clothes hanging up, trapped inside

mosquito nets etc. were aspirated and blown into a holding cup for identification and other

analyses such as further speciation, and susceptibility testing etc.



3.1.6 Preservation of Mosquitoes All Anopheles mosquito specimens from resistance tests, HRC and HLP were preserved for

further analysis (see below) in an entomology laboratory at the Liverpool School of Tropical

Medicine (LSTM).

Culicine species either captured in HLC, HRC or emerging from captured larvae were also

preserved for precise determination of species at The Natural History Museum in London,

UK. Mosquitoes were placed individually in labelled 0.5ml Eppendorf tubes. The tubes were

colour coded based on the sampling method and those that survivors and died in the fields

resistance tests. Tubes were individually pierced and kept in sealed plastic bags with silica

gel to ensure the mosquitoes remained dry until further analysis.

3.2 Laboratory techniques The analysis of the mosquito samples is being conducted in collaboration with the Liverpool

University School of Tropical Medicine. Due to technical difficulties and capacity at the

laboratory the final analysis of the samples has not been completed as yet. These results will

be added to the report as a supplement when available.

There is scarce information on vector species composition in Guinea. Previous studies have

reported Anopheles gambiae s.l. (77%) as the most common Anopheles collected in four

different regions with An. funestus contributing 20%. The specimens of the An. gambiae

complex were reported as predominantly An. gambiae S form (97.6%) with 1.4% of An.

gambiae M form found in Kérouané only, and 1% of An. arabiensis which was present in all

four study sites. Anopheles gambiae S form and An. funestus were found to be infected with

P. falciparum, with infectivity rates of 4.1% and 4.4% and inoculation rates of 0.60 and 0.19

infected bite/person/night, respectively. In addition, a high level (79%) of the knockdown

resistance (kdr) L1014F mutation was reported in the populations of An. gambiae S form

[17].

There have been no studies reported from the areas sampled in this study and the

implications of the above reported data are significant in terms of epidemiology and control of

malaria vectors concerned. It is therefore crucial to accurately determine species

composition, sporozoite rates and to elucidate any resistance mechanism occurring.



3.2.1 Species Identification Anopheles gambiae s.s. occurs as two, often-sympatric races, termed the M and S molecular

forms. Clarifying the genetic population structure of the An. gambiae s.s. complex is critical

for determining which genetic units of the complex are the main vectors of malaria. This will

assist in supporting the understanding of the ecological and ethological differences relevant

to disease transmission and epidemiology in the area and assist in the design and

implementation of appropriate vector control strategies.

An example of the relevance of this information to control can be seen in Burkina Faso where

the M molecular form shows the closest association with the domestic environment and

larval habitats created by human activities, whereas the S form is more frequent in rain-

dependent temporary breeding sites. This has serious consequences for malaria

transmission as in dry areas of West Africa the M form is able to exploit breeding

opportunities due to human activities where in the past only the far less anthropophilic An.

arabiensis would have been found [18].

3.2.2 Detection of knockdown resistance gene Knock-down resistance (kdr) to organochloride and pyrethroid classes of insecticide in

Anopheles gambiae s.s (as the main vector species) is important to analyse, as this can

influence decisions related to control and choice of insecticide. Kdr is two alternative point

mutations which can lead to resistance to these classes of insecticide and render control

programmes less effective.

In An. gambiae S-form populations, the mutation appears to be widespread in West Africa

and has been reported from Uganda, Kenya, Gabon, Cameroon and Equatorial Guinea. In

M-form populations these mutations have been found, although it is less widespread, and at

lower frequencies than in sympatric S-form populations [19].



3.2.3 Detection of Sporozoites of Plasmodium falciparum The detection of the sporozoite rates in mosquitoes is a reflection of the fraction of the vector

mosquitoes that are infectious and can thus transmit infection. This was important at the local

level as it supports a better understanding of the dynamics of malaria transmission and helps

to define when and where the greatest risk occurs and facilitates the development of

appropriate control strategies. It can also be used to determine which vector species

contribute most to malaria transmission which is a function of the vector population density

and sporozoite rate.

The head and thorax of the mosquito were sampled and tested using a polymerase chain

reaction technique to determine the presence of sporozoites in the salivary glands of the

mosquito [22]. This was performed to determine how the level of risk of malaria transmission

within an area may compare with other (or surrounding) populations, which will help identify

key differences/similarities and highlight corresponding risk factors. This was especially

important considering the different ecological and urban/rural areas in the study area to

determine if the risk for transmission may differ in these areas.

The parasite prevalence survey will be used as a measure of local transmission but if the

Project expands and a formal vector control team is established which is supported by skilled

staff then it may be worthwhile tracking entomological inoculation rates, or human biting

rates, i.e. the number of bites per person per day from an infected mosquito.



4 Results 4.1 Morphological Identification of Mosquitoes Captured in Field Morphological identification of mosquitoes either reared from larvae or captured in HLC or

HRC was carried out as far as possible in the field. However, the sibling species of the

Anopheles gambiae species complex cannot be identified morphologically in field. It was

considered important to determine the composition of the species complex as their

behaviours differ greatly in their breeding site preferences, host blood meal preferences as

well as resting sites. These can all influence decision making regarding control programmes.

Species which were identified in the field are described in Table 6 with some remaining

Anopheles gambiae species complex still being analysed at the laboratory. All anomalous

larvae and pupae are in the process of identification with the Natural History Museum.

Table 6: Mosquito species found in the field

Genus Species Sampling method Sentinel site Anopheles gambiae s.l.* HLC, HRC Manké, Maférinyah Anopheles funestus HLC Manké Culex quinquefasciatus HLC, HRC Manké Aedes aegypti HLC (day) Manké

*it is possible that these include An. gambiae s.s., An. arabiensis and An. melas

4.1.1 Human Landing Catches In Maférinyah, one human landing catch was done at Fanye. However, despite the large

breeding site nearby only one mosquito was caught by 4 catchers from 6pm to 6am. This

was probably due to the wet and windy weather conditions during the evening. The time

scheduled for this activity was limited and when rescheduled the weather was similar and the

HLC was abandoned. The mosquito caught was identified as Anopheles gambiae s.l.

In Manké, the HLC yielded large numbers of anthropophagic mosquitoes despite severe

thunderstorms on one night. The mosquitoes were so voracious that they were prevented

from flying and biting only during the height of the rainfall. Mosquitoes caught were identified

as Anopheles gambiae s.l., Anopheles funestus and Culex quinquefasciatus. At Manké it

was apparent that the houses closer to the breeding site were subject to more intense biting

at night.



4.1.2 House Resting Catches House resting catches were carried out at Manké, Maférinyah and Yindi-Famoriah.

Mosquitoes were caught at Manké and Maférinyah although the yields were small compared

to the density of larvae in nearby breeding sites and the high yields of mosquitoes caught

during the HLC at Manké.

The species caught were Anopheles gambiae s.l. (both sites) and Culex quinquefasciatus

(Manké only). The laboratory analysis of species may guide speculation as to the reason for

the low indoor resting catches compared to high indoor biting.

4.2 Morphological Identification of Larvae or Adult Mosquitoes

Reared from Larvae Larvae which emerged into adults and were identified morphologically in the field are shown

below in Table 7.

Table 7: Larvae species found in the field

Genus Species Breeding site type Sentinel site

Anopheles gambiae s.l.* Rice field / marsh / chilli farm

Manké, Matakang Maférinyah

Anopheles funestus Rice field / marsh Manké, Matakang Anopheles coustani Marsh Manké Culex quinquefasciatus Rice field Manké Toxorhyncites - Rice field Manké

*It is possible that these include An. gambiae s.s., An. arabiensis and An. melas



4.3 Resistance Tests Anopheles gambiae s.l., which is the principal malaria vector in Guinea, was exposed to the

insecticide treated papers as per the approach described in Appendix 1.

Table 8 described the levels defined by the WHO to determine the susceptibility and

probability of resistance based on exposure to the different insecticide treated papers.

Table 8: Susceptibility/Resistance levels according to WHO

Mortality Level of susceptibility

98-100% Susceptible

80-97%1,2 Possibility of resistance

<80%2 Resistance 1 Resistance that needs to be confirmed by testing for resistance mechanisms such as kdr 2 Where mortality of <95% occurs in tests that have been conducted under optimal conditions with a

sample size of more than 100 mosquitoes, resistance is strongly suspected.

The results from exposing adult Anopheles gambiae s.l. mosquitoes to the insecticide

impregnated papers for one hour and then holding for 24 hours with no further exposure are

shown below in Table 9. Susceptibility is corrected for control mortality using Abbott’s

formula as described in appendix 1.

Table 9: Susceptibility of An. gambiae s.l.

Insecticide used (concentration), Insecticide class

Ile Kaback Forécariah Mainland

Manké Matakang Maférinyah Yindi Deltamethrin (0.05%), Pyrethroid 100% 92%* 87%* -

Bendiocarb (0.1%), Carbamate 100% 90%* 95%* -

Fenitrothion (1.0%) Organophosphate 100% 97%* 100% -

*kdr-testing underway

From these results it appears that at Manké there is no indication of resistance to any of the

three classes of insecticide tested, while at Maférinyah and Matakang further evaluation

should be carried out to clarify possible resistance to pyrethroid class of insecticides and the

carbamate class of insecticides. The resistance tests using mosquitoes from Matakang

exposed to Fenitrothion was at 97%, suggesting possible resistance. Further testing of more



mosquitoes may confirm whether true resistance exists. The Maférinyah mosquitoes were

fully susceptible to organophosphates (i.e. Fenitrothion).

As a limitation to the present study, the numbers of Anopheles funestus larvae were

insufficient at the time of the study to do any significant resistance testing. But nonetheless,

adult An. funestus are efficient vectors of malaria and may contribute considerably to malaria

transmission at different times of the year, and may have different resistance profiles.

4.4 Laboratory Assays The laboratory assays conducted so far are on mosquitoes reared from larvae at Manké.

Speciation of 100 mosquitoes so far has shown all to be Anopheles gambiae M form. The M

molecular form has only been reported previously at Kérouané, however, even there it

contributed only 1.4% to the An. gambiae population. Assays are on-going and will be

reported on when completed. Since M and S forms differ in their behaviour and breeding

habits, the assay results will be critical to understanding malaria epidemiology in this area

and optimising control recommendations.



5 Summary Discussion on Key Findings and Recommendations

5.1 Malaria and Associated Vectors 5.1.1 Ecology and Potential Vectors The ecology in the area is suitable for the two most efficient vectors in sub-Saharan Africa,

namely, An. gambiae and An. funestus. The rice field and general habitats in the area are

well suited to the proliferation of these efficient vectors. While An. gambiae was the

predominant species found during the assessment there is the potential that An. funestus

may dominate at different times of the year. This may sustain transmission patterns from one

season to the next and also pose challenges to control measures [23]. It will thus be

important to develop an entomological surveillance programme as part of the Project vector

control programme to effectively manage control activities based on the predominant

circulating vectors.

The rice fields create the dominant habitat in the area but as the survey was conducted when

heavy rains fell this may change as the available breeding sites transform. It is likely that An.

gambiae will predominate during construction of the proposed port facilities due to the

environmental modifications and these vectors preferred breeding habits. It will thus be

essential to manage source reduction aggressively and perform IRS as the main vector

control activity.

In the dry season there is the potential that An. arabiensis could emerge as an important

local vector. This species prefers lower humidity so the dry harmattan winds that blow from

the Sahara during the winter months from November to March. This is a theoretical risk and

of probably lower significance than the seasonal risk posed by An. funestus.

An. melas is associated with the saline water conditions and the mangrove swamps but was

not isolated in the assessment. The role that this species may play in disease transmission

will require on-going surveillance, especially with the potential for the Project development to

alter the local environment with dredging and construction along the banks of the Morebaya

River.



5.1.2 Species Composition and Behaviour The key findings of the assessment based on areas of activity are summarized below:

Larval sampling:

• In Manké, adult mosquitoes collected were located in a number of locations within

the rice field growing area. The larvae collected must have been from a large number

of adult females since they were collected at many different locations considerable

distances from each other. While these numerous breeding sites outline the general

susceptibility of the local population it was relevant to note the rice fields closest to

the village had the highest larval sampling yields.

• In Matakang, the adult mosquitoes used were also from a number of locations within

a rice field covering an area of approximately 20 hectares. Larvae were found in a

variety of locations within the rice field at considerable distances from each other so

we can again conclude that they represent the offspring of a large number of adult

female mosquitoes and are representative of the local population. The rice field was

located adjacent to a Chinese camp and on the edge of the beach close to the sea. It

is possible that Anopheles melas form a proportion of this population since the close

proximity to the sea is likely to cause brackish water at this site. The species assays

being conducted will answer this query and may explain the difference in resistance

results over such a short distance from Manké.

• In Maférinyah, the larvae were collected from a number of sites. The most prolific

site was a duck pond at Fanye. There was such a high density of larvae here that

they must have come from a large number of adult female mosquitoes. However, the

population may be highly localized as the HLC conducted approximately in 300

meters distance from the breeding site yielded only a single mosquito which was a

surprise.

• In Yindi-Famoriah, since no Anopheles larvae were found, this site should be

elucidated at a time of year appropriate to mosquito breeding.

Human landing and House resting catches:

The human landing catches conducted at Manké showed the Anopheles mosquitoes caught

to be at least as exophagic (outdoor feeding) as endophagic (indoor feeding). Further

information assessing the endophilic and exophilic behaviour of the vector populations at the

four sites should be obtained prior to considering a control programme based on endophagic

behaviour such as ITNs or based on endophily such as IRS. For the present time it is

recommended that an integrated approach be adopted using IRS, ITNs and larvaciding as



key vector control activities and source reduction as a primary objective (not on agricultural

sources such as rice fields etc). Different classes of insecticides should be used for

larvaciding and for adult control, where this is possible.

House resting catches were only successful in Manké where high numbers of both

Anopheles and Culex were caught throughout the night from 6pm to 6am. Both, Anopheles

and Culex species were found resting indoors but in much lower numbers than was expected

considering the high numbers caught in HLC.

Only small numbers of An. funestus were found during the period of study. An. funestus is

often an important vector of malaria during certain times of the year, especially at the end of

the rainy season when it frequently takes over from An. gambiae as the principal vector

significantly extending the malaria transmission season. The importance of this vector should

not be underestimated and should be fully investigated as part of any control programmes.

Surveillance of this species is essential to ensure effective control and to manage risks. An.

funestus is extremely difficult to breed in captivity but rearing of adults from larvae for

resistance testing is possible as well as assessing behavioural characteristics of the natural

population. This should be undertaken during different seasons of the year so that the

contribution to overall transmission can be assessed and appropriate control measures for

this vector initiated.

5.1.3 Resistance Profile Although the sampling sites chosen were within the same sub-prefecture, considerable

variation in resistance profiles was seen on field analysis. While the kdr testing is still on-

going, it may be useful to support additional assessments on species composition and

resistance patterns as part of the surveillance activities of any planned vector control

activities. In the interim, it is recommended to use an organophosphate class of insecticide

for IRS in the first instance until the status of the pyrethroid and carbamate class is defined.

In any event it is recommended to adopt a mosaic approach on the choice of insecticide,

alternating between the different the three classes.

Organophosphates, in the form of pirimiphos-methyl (Vectoguard 40WP® or Actellic 50 EC),

is recommended, but this will require a full health, safety and environmental review prior to

use. There is a newer Actellic formulation which is increasing the residual life of the

compound but this is not yet commercially available. Alternatively, compounds with



Fenitrothion as the active formulation can be used as these have a longer residual life. Spray

operators will need to be enrolled in the biological monitoring programme as part of the

workplace occupational health programme as a similar exposure group to

organophosphates.

The recommended pyrethroid can be in the form of micro-encapsulated lamda-cyhalothrin

(Icon 10CS®) or deltamethrin (K-othrine WG 250). If LLINs are used extensively then

another class of insecticide should be used in the early stages of the IRS programme- it is

recommended to use a Carbamate in this instance.

Larvaciding should be considered in the integrated approach especially in areas where active

An. funestus breeding is found. Ideally a biological larvacide should be used or a form of

larvacide oil. This approach will reduce the potential development of resistance due to

organophosphates been used on both, larvae and adults.

Space spraying should be limited based on entomological surveillance to target a specific

reduction in vector density and also for outbreaks of other diseases. Ideally the Project

should have the capability to undertaken space spray with either a pyrethroid or

organophosphate based insecticide especially in preparation for Aedes population being

detected and requiring urgent control.

5.2 Other Vectors of Medical Importance 5.2.1 Ecology and Potential Vectors The environment was conducive to the Aedes and Culex group of mosquitoes. It is likely that

that these species will circulate through the year. Aedes was of particular concern due to

potential risks from YF, dengue and Chikungunya fever, especially with the establishment of

the Port and rail line to the interior of the country.

5.2.2 Species Composition and Behaviour During the period of study only one adult Aedes aegypti was found feeding during the day (in

Manké). However questioning of the communities living at Manké and also at Maférinyah

suggested that Ae. aegypti can be found in considerable numbers at different times of the

year (with reported high number of a vicious day biting black and white stripey mosquito in

community consultations).



It would be expedient to set up monitoring of day biting Aedes mosquitoes as part of any

proposed control programmes so that seasonal patterns can be determined and risk period

identified. Control programmes should be focussed on source reduction in the proposed work

camps and communities and larvaciding in the workplace and camps. Larvaciding should be

reserved for areas where source reduction is a challenge (for example lay down yards, on

top of containers and in tyres). This should be performed with Temephos (Abate® 1SG (1%

sand granule) or Abate 5PG (5% plaster pellets)) applied as granules or a liquid formulation.

Temephos is an organophosphate based product.

A number of Culicine species were found, these are in the process of identification, however

they are unlikely to be of importance in disease transmission.

Tsetse, blackflies and ticks were not trapped during this study. Since they are also vectors of

medical importance they should be assessed as part of any surveillance programme. Tsetse

populations especially should be evaluated and monitored regularly.



6 References 1. SHAPE Consulting Limited, Health Impact Assessment: Baseline Health Survey -

Simandou Project, Port Development Area, Rio Tinto Iron Ore Simfer SA, Guinea, October 2011.

2. Newfields LLC, Health Impact Assessment: Scoping Study - Simandou Project, Port Development Area, Rio Tinto Iron Ore Simfer SA, Guinea, September 2010.

3. Jentes, E.S., et al., Acute arboviral infections in Guinea, West Africa, 2006. Am J Trop Med Hyg, 2010. 83(2): p. 388-94.

4. WHO. Global Altert and Response - Guinea. 2012a [cited March 2012; Available from: http://www.who.int/csr/don/archive/country/gin/en/.

5. Gratz, N.G. and A.B. Knudsen. Dengue Bulletin Volume 21: The Rise and Spread of Dengue, Dengue Haemorrhagic Fever and its Vectors 19950-1990. 1997; Available from: http://www.searo.who.int/en/section10/section332/section519_2401.htm.

6. WHO. Other arbo-viral diseases: Chikungunya. 2012b [cited March 2012; Available from: http://www.who.int/denguecontrol/arbo-viral/other_arboviral_chikungunya/en/index.html.

7. de Souza, D., et al., Environmental factors associated with the distribution of Anopheles gambiae s.s in Ghana; an important vector of lymphatic filariasis and malaria. PLoS One, 2010. 5(3): p. e9927.

8. Konstantinov, O.K., et al., [African trypanosomiasis in the Republic of Guinea]. Med Parazitol (Mosk), 2008(3): p. 36-40.

9. Ilboudo, H., Jamonneau,V.,Camara,M.,Camara,O.,Dama,E.,Leno,M.,Ouendeno,F.,Courtin,F.,Sakande,H.,Sanon,R.,Kabore,J.,Coulibaly,B.,N'Dri,L.,Diarra,A.,N'Goran,E.,Bucheton,B., Diversity of resonse to Trypanosoma brucei gambiense infections in the Foerecariah mangrove focus (Guinea): perspectives for a better control of sleeping sickness. Microbes and Infection, 2011. 13(11): p. 943-952.

10. Guillet, P., et al., Impact of combined large-scale ivermectin distribution and vector control on transmission of Onchocerca volvulus in the Niger basin, Guinea. Bull World Health Organ, 1995. 73(2): p. 199-205.

11. Boakye, D.A., Back,C.,Fiasorgbor,G.K.,Sib,A.P.,Coulibaly,Y., Sibling species dostributions of the Simulium damnosum complex in the west African Onchocerciasis Control Programme area during the decade 1984-93, following intensive larviciding since 1974. Med Vet Entomol., 1998. 12(4): p. 345-58.

12. Ergonul, O., Crimean-Congo haemorrhagic fever. Lancet Infect Dis, 2006. 6(4): p. 203-14.

13. Health, T.C.f.F.S.a.P., Crimean-Congo hemorrhagic fever, 2009. 14. Parola, P.,

Diatta,G.,Socolovschi,C.,Mediannikov,O.,Tall,A.,Bassene,H.,Trape,J.F.,Raoult,D., Tick-borne relapsing fever Borreliosis,rural Senegal. Emerging Infectious Diseases, 2011. 17(5): p. 883-885.

15. Mediannikov, O., Diatta,G.,Zolia,Y.,Balde,M.C.,Kohar,H.,Trape,J.F.,Raoult,D., Tick-borne rickettsiae in Guinea and Liberia. Ticks Tick Borne Dis., 2012. 3(1): p. 43-8.

16. Food and Agricultural Organisaton,



17. Carnevale, P., Toto,J.C.,Guibert,P.,Keita,M.,Manguin,S., Entomological survey and report of a knockdown resistance mutation in the malaria vector Anopheles gambiae from the Republic of Guinea. Trans.Roy.Soc.Trop.Med.Hyg., 2010. 104(7): p. 484-489.

18. Caputo, B., Santolamazza,F.,Vicente,J.L.,Nwakanma,D.C.,Jawara,M.,Palsson,K.,Jaenson,T.,White,B.J.,Mancini,E.,Petrarca,V.,Conway,D.J.,Besansky,N.J.,Pinto,J.,della Torre,A., The "Far-West" of Anopheles gambiae molecular forms. PLoS one, 2011. 6(2): p. e16415.doi:10.1371/journal.pone.0016415.

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7 Appendices 7.1 Appendix 1: Standard Operating Procedure (SOP), WHO 1998 –

Resistance/Susceptibility Test for Anopheles mosquitoes A standard diagnostic test kit was used comprising of 14 plastic tubes (125 mm in length and

44 mm in diameter), with each tube fitted at one end with 16-mesh screen. The twelve tubes

include:

• Six tubes marked with a red dot for use as exposure tubes i.e. for exposing

mosquitoes to the insecticide impregnated papers.

• Two tubes marked with a green dot for use as control tubes i.e. for exposure of

mosquitoes to the oil treated control papers without insecticide.

• Six tubes with a green dot for use as holding tubes i.e. for pre-test sorting and post-

exposure observation.

• Six slide units, each with a screw cap on either side and provided with a 20mm filling

hole.

• 100 sheets of clean paper (12cm x 15cm) for lining the holding tubes.

• 14 spring wire clips to hold the papers in position against the walls of the tubes. Of

these 8 steel clips are used in the 7 holding tubes and the 1 control exposure tubes

and 6 copper clips are used in the insecticide exposure tubes.

• Two aspirators of a diameter that can pass snugly through the 20mm filling hole of

the slide unit.

• One roll adhesive tape.

• Instruction sheet.

• Report forms.

• Insecticide impregnated papers.

The insecticide impregnated papers used in standard resistance/susceptibility test are

prepared at University Sains Malaysia, Penang, Malaysia on behalf of WHO. The papers are

prepared with a discriminating dose of insecticide i.e. a dose which should allow a decision to

be made on the resistance/susceptibility of the population being tested depending on the

survival rate at 24 hours of the mosquitoes after exposure to the insecticide for one hour.

simandou project port development area · 2018-11-21 · 1.1.2 lymphatic filariasis ... sop...

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