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Argulus canadensis is a crustacean ectoparasite observed increasingly on wild migrating adult Atlantic salmon. We investigated temperature and salinity tolerance regarding development, survival and hatch of A. canadensis eggs to help understand spatiotemporal features of transmission. Argulus canadensis eggs differentiate to pharate embryos by 35 days buttheir hatch is protracted to ~7 months. Cold treatment ⩾75 days mimics overwintering and terminates egg diapause, with 84.6% (72.1-100%) metanauplius hatch induced ⩾13 °C and synchronized to 3-4 weeks. Inter- and intra-clutch variability and protracted hatch in the absence of cold-temperature termination of diapause is compatible with bet hedging. Whereas diapause likely promotes phenological synchrony for host colocalization, bet hedging could afford temporal plasticity to promote host encounter during environmental change. Our egg storage and hatch induction/synchronization methodologies can be exploited for empirical investigations. Salinity tolerance reveals both significantly higher embryonic development (94.4 ± 3.5% vs 61.7 ± 24.6%) and metanauplius hatch (53.3 ± 7.5% vs 10.1 ± 8.2%) for eggs in freshwater than at 17 ppt. Unhatched embryos were alive in freshwater by the end of the trial (213 days) but were dead/dying at 17 ppt. Eggs did not develop at 34 ppt. Salinity tolerance of A. canadensis eggs supports riverine transmission to adult Atlantic salmon during return to freshwater for mating each year.

With expanding human populations, the food sector has faced constant pressure to sustainably expand and meet global production demands. In aquaculture this frequently manifests in an animal welfare crisis, with fish increasingly farmed under high production, high stress conditions. These intense environments can result in fish stocks having a high susceptibility to infection, with parasites and associated disease one of the main factors limiting industry growth. Prediction of infection dynamics is key to preventative treatment and mitigation. Considering the climatic and technology driven changes facing aquaculture, an understanding of how parasites react across a spectrum of conditions is required. Here we assessed the impact of temperature, infection density and host species on the life history traits of Argulus foliaceus, a common palearctic fish louse, representative of a parasite group problematic in freshwater aquaculture and fisheries worldwide. Temperature significantly affected development, growth and survival; parasites hatched and developed faster at higher temperatures, but also experienced shorter lifespans when maintained off the host. At high temperatures, these parasites will likely experience a short generation time as their life history traits are completed more rapidly. A. foliaceus additionally grew faster on natural hosts and at lower infection densities. Ultimately such results contribute to prediction of population dynamics, aiding development of effective control to improve animal welfare and reduce industry loss., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

The knowledge of mucosa-associated molecular events that occur during infections is scarce despite the well-established importance of mucus in fish immunity. Using qRT-PCR, we analyzed the immune gene expression patterns in mucus of Labeo rohita experimentally infected with an ectoparasite Argulus siamensis. Mucus samples were collected at 0 h, 12 h, 24 h, 3 d, 7 d, 15 d, and 30 d post challenge of L. rohita with metanauplii of A. siamensis. All interleukins studied herein (IL 6, IL 15, and IL 1β) showed significant upregulation of expression levels in mucus of A. siamensis-infected fish compared to control samples. Further, the expression levels of molecules involved in pathogen recognition, toll like receptor 22, and pathogen presentation, β2 microglobulin, were found to be significantly upregulated in experimental samples until 7 d post challenge compared to control samples. The upregulated expression of lysozyme G at all time points post infection indicated the early activation of acute phase responses in mucus of infected L. rohita. Moreover, the expression levels of natural killer cell enhancing factor B were found to be higher in infected fish than they were in the control fish. The early upregulation of the immune genes observed herein reinforces the role of mucus as the first line of defense against pathogenic assault; furthermore, it expands our understanding of mucosal-immune responses to A. siamensis infection, which can aid development of immunological interventions.

We have created barcode library for common Argulus spp. infecting Carassius auratus, which could also be used to identify premature forms of Argulus spp. even by non-professionals. Infected C. auratus was examined and purchased from ornamental fish-trading centers and the adult life stage of Argulus spp. was identified and DNA barcoded. The eggs of Argulus spp. were collected using bottle implants. The collected eggs are barcoded and precisely identified by matching with the adult sequences. Four species of adult Argulus spp. were identified, namely Argulus japonicus, Argulus indicus, Argulus siamensis, and Argulus foliaceus. Precise identification of egg samples was done by two different analyses, namely (i) BLAST analysis and (ii) phylogenetic clustering of adults and eggs. All egg samples including the control were precisely identified by BLAST analysis and the results are consistent with phylogenetic clustering of adult and egg's DNA barcodes. In order to establish the DNA barcode technology for the identification of all Argulus spp and its premature forms, the development of full-fledged barcode library that includes all species of this genus is very important for the benefit of ornamental fish industries.

Fish lice (Argulus spp.) are obligate ectoparasites, which contrary to most aquatic parasites, retain the ability to swim freely throughout the whole of their life. In fish farms, they can quickly increase in numbers and without effective control cause argulosis, which results in the reduced growth and survival of their fish hosts. The morphology of Argulus spp, including their sensory organs, is suitable for both parasitism and free-swimming. By spending a considerable amount of time away from their host, these parasites risk being excessively dispersed, which could endanger mating success. Here we present a review of recent studies on the behaviour of Argulus spp, especially the aggregative behaviour that mitigates the dilution of the parasite population. Aggregation of parasites, which is especially important during the period of reproduction, occurs on different scales and involves both the aggregation of the host and the aggregation of the parasites on the host. The main behavioural adaptations of Argulus spp, including searches for hosts and mates, host manipulation and host choice, are all focused on the fish. As these ectoparasites repeatedly change hosts and inflict skin damage, they can act as vectors for fish pathogens. The development of environmentally friendly measures for the control and prevention of argulosis needs to take into account the behaviour of the parasites.

The present investigation was carried out to underscore the developmental events of a crustacean ectoparasite of fish, Argulus bengalensis. Serial histological sections of the embryo were made at lateral, sagittal and longitudinal planes to explain its cleavage, gastrulation and axis specification. The centrolecithal egg of A. bengalensis underwent meroblastic superficial cleavage. The cleavage initiated at the future dorsal side of the egg within 5 h to 5 h and 30 min of incubation. Consequently, a small mass of energids appeared superficially at the future dorsal side within 6 h. Later, energids were found at the future ventral and lateral sides. A syncytial blastoderm was formed around the centrally placed yolk material which was transformed into a cellular blastoderm within 30 h of incubation. In the blastoderm, two cell masses were formed at the dorsal and ventral part which initially extended towards each other and later spread out though future anterior-posterior direction. The pressure exerted by the cell flow displaced the entire yolk material at the future postero-ventral side. At the time of egg laying, a prototype of the embryonic axes is determined. The substratum side of the egg formed the dorsal part, whilst the side facing water turned to the ventral part. The broader end of the egg formed the anterior side and the narrow end formed the posterior side of the embryo. The anterior-posterior axis formation was initiated within 72 to 96 h of incubation when the blastodermal cells displaced the yolk material at the future posterior end. Within 120 h of incubation, the germ layers of the embryo were determined. The study reveals that the cleavage pattern of A. bengalensis shows close similarities with that of the Malacostraca amongst the crustaceans and dipteran and hymenopteran amongst the insects.

To assess the effects of abiotic factors on Argulus japonicus occurrence in freshwater ecosystem, we sampled fish, free-swimming parasites, and submerged objects from rivers, fish farms, and ponds (reservoirs and lakes) at 27 locations in Guangdong province, China, from July, 2010, to March, 2013. Friedman's test and Spearman's correlation coefficient (r) were used to investigate relationship between A. japonicus occurrence and abiotic factors in three aquatic systems. Correlation of monthly density, mean density, prevalence, abundance, and intensity of A. japonicus with water temperature, biochemical oxygen demand of 5 days, chemical oxygen demand, and ammonia were very significant, but negative relation with pH, dissolved oxygen, and total phosphorus components. Nevertheless, total nitrogen components showed an insignificant impact on A. japonicus incidence. In addition, water temperature and stability were two key factors controlling A. japonicus infestation. Our study revealed the capability of A. japonicus to inhabit and prevail in heavily polluted freshwater. Results demonstrate that A. japonicus has good adaptability to the environmental factors' stress but cannot be used as a biological indicator of environmental pollution.

Recent advances in high-throughput sequencing have facilitated the generation of large transcriptomic datasets for an ever-growing number of crustaceans, one being the carp louse Argulus siamensis. This and other members of the subclass Branchiura are obligate fish ectoparasites, and as such, are a major concern for commercial aquaculture. Using the extant transcriptome shotgun assembly (TSA) sequences for A. siamensis, 27 transcripts encoding putative neuropeptide precursors were identified, and their pre/preprohormones deduced and characterized using a well-established bioinformatics workflow. The structures of 105 distinct peptides were predicted from the deduced proteins, including isoforms of adipokinetic hormone (AKH), allatostatin A, allatostatin B, allatostatin C, allatotropin, bursicon α, bursicon β, crustacean cardioactive peptide (CCAP), diuretic hormone 31, diuretic hormone 44, eclosion hormone, myosuppressin, neuroparsin, neuropeptide Y, orcokinin, pigment dispersing hormone, proctolin, short neuropeptide F, SIFamide, sulfakinin and tachykinin-related peptide. While several of the predicted peptides are known from other crustacean and/or insect species, e.g. RYLPT, a broadly conserved arthropod proctolin isoform, and PFCNAFTGCamide (disulfide bridging between the two cysteines), the stereotypical crustacean CCAP, the vast majority of them are described here for the first time, e.g. pQVNFSTKWamide, a new AKH/red pigment concentrating hormone superfamily member, pQEGLDHMFMRFamide, a novel myosuppressin, and SYKSKPPFNGSIFamide, a new member of the SIFamide family. As the peptides presented here are the only ones thus far described from A. siamensis, or for that matter, any branchiuran, they represent a new resource to begin investigations of peptidergic control of physiology and behavior in this and other related aquacultural pests., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Global climate change (GCC) is expected to affect key environmental variables such as temperature and rainfall, which in turn influence the infection dynamics of metazoan parasites in tropical aquatic hosts. Thus, our aim was to determine how temporal patterns of temperature and rainfall influence the mean abundance and aggregation of three parasite species of the fish Cichlasoma urophthalmus from Yucatán, México. We calculated mean abundance and the aggregation parameter of the negative binomial distribution k for the larval digeneans Oligogonotylus manteri and Ascocotyle (Phagicola) nana and the ectoparasite Argulus yucatanus monthly from April 2005 to December 2010. Fourier analysis of time series and cross-correlations were used to determine potential associations between mean abundance and k for the three parasite species with water temperature and rainfall. Both O. manteri and A. (Ph.) nana exhibited their highest frequency peaks in mean abundance at 6 and 12 months, respectively, while their peak in k occurred every 24 months. For A. yucatanus the frequency peaks in mean abundance and k occurred every 12 months. We suggest that the level of aggregation at 24 months of O. manteri increases the likelihood of fish mortality. Such a scenario is less likely for A. (Ph.) nana and A. yucatanus, due to their low infection levels. Our findings suggest that under the conditions of GCC it would be reasonable to expect higher levels of parasite aggregation in tropical aquatic hosts, in turn leading to a potential increase in parasite-induced host mortality.

Parasite intensity dynamics can be strongly affected by ecosystem abiotic components. We conducted this study to investigate how the intensity of Argulus sp. is influenced by physicochemical factors in three water bodies - river, fish farm, and reservoir - and to find the occurrence of Argulus sp. under different conditions. Argulus sp. specimens were collected using different methods - gill-netting 30 × 1.5 m, 40 mm mesh monofilament net and set from a small boat, beach seining and plankton net - from July 2010 until Jun 2011 at Guangdong province, China. Main water parameters taken for the assessment consisting of temperature, pH, DO, biochemical oxygen demand of 5 days (BOD5), chemical oxygen demand (COD), NH (4) (+) , total nitrogen (TN) and total phosphorus (TP). The results show that the ratio of female to male Argulus sp. during the studied period was 2.3:1. The presence and intensity of Argulus sp. were temperature-dependent and variable; pH values were lower than standers with high intensity of Argulus sp. Argulus sp. has the ability to live in low dissolved oxygen (DO) concentration. Relatively high concentrations of BOD5, COD, NH (4) (+) , TN and TP were detected with related variable intensity of Argulus sp., which is supposed to be a freshwater fish ectoparasite found in polluted ecosystems. It is concluded that the fish ectoparasite Argulus sp. have a strong potency of genetic adaptation impacted by environmental factors. However, this needs further study in order to gain insight into the question of the likelihood of adaptation to abiotic factors' variation.

Eighteen female specimens of Argulus vittatus (Rafinesque-Smaltz, 1814) were recently collected from the Algerian coast. As until now this species was poorly described, this contribution redescribes this species with more precise drawings on the general morphology and appendages, using this fresh material. For the first time, two new hosts are identified. Host specificity and some ecological data are also reported.

The growth kinetics of aquatic worms was investigated from juvenile to decline phase for 18 weeks by cultivating with activated sludge in batch test. Results showed that the growth of aquatic worms well fit Gauss function for cultivating 18 weeks. The maximum specific growth rate, growth yield of aquatic worms and sludge reduction rate was 0.41 d(-1), 0.32 and 25.5%, respectively. When the concentration of substrate and dissolved oxygen change from low to high, the relationship between the specific growth rate with dissolved oxygen, and substrate concentration meet the Monod equation. Compared with the dissolved oxygen, the substrate concentration had greater effect on the specific growth rate of aquatic worms, and aquatic worms can live in the environment with low dissolved oxygen. Furthermore, the breath test showed the oxygen uptake rate of aquatic worms was almost 6.39, 10.10, 11.31 and 5.74 mg x (L x g x h)(-1) from juvenile to decline phase, the dissolved oxygen demand of the rapid growth and mature stage was higher than juvenile and decline phase.

In this study we examine differences in the occurrence of life history stages of the destructive fish ectoparasite Argulus foliaceus (L., 1758) on eight fish species (stickleback, rudd, roach, gudgeon, bream, tench, crucian carp and common carp) sampled from a mixed-species recreational fishing lake on nine occasions during late spring and summer. Total numbers ofA. foliaceus, as well as the number of larval, juvenile and adult parasite stages, from each fish were recorded along with the fish species. Lice generally exhibited an aggregated distribution approximating a negative binomial distribution. Significant differences in the prevalence, intensity and intensity frequency distribution were observed between life history stages and between host species. In general, all life history stages of A. foliaceus exhibited an over-dispersed distribution. However, larval lice did show some degree of aggregation particularly within the stickleback samples. Infection data for parasite larval stages suggested that sticklebacks are more likely to be infected than other host species. For adult lice, however, carp appeared to be the main host. We propose that A. foliaceus infection characteristics are predominantly determined by the level of host exposure to the parasite and its life history stages (larval, juvenile and adult) rather than by an innate difference in host susceptibility related to individual host factors such as immune responses. We conclude that host exposure is determined by the parasite-host behavioural interplay related to species-specific ecology and behavioural traits such as microhabitat preference and normal swimming speed.

We tested the hypothesis that host specificity in ectoparasites does not depend exclusively on the features of the host but also on surrounding habitats, using 2 fish ectoparasites, Argulus coregoni and A. foliaceus (Crustacea: Branchiura), occurring sympatrically in Finnish lakes. Although these parasites are considered to be of low specificity, we found that the larger of the 2 species, A. coregoni developed a pronounced preference for salmonid hosts at the beginning of maturation (defined by the presence of copulating specimens). Argulus foliaceus infects a much wider range of fish hosts. We showed that specialization of A. coregoni on salmonids does not necessarily result from incompatibility with other fishes, but could instead reflect higher sensitivity of oxygen depletion compared with A. foliaceus. Adult A. coregoni may meet these demands by attaching to salmonids, the typical inhabitants of well-aerated waters. Young parasites of both species showed little host specificity and attached mainly to fishes with higher body reflectivity. In host choice experiments, A. coregoni of 4-5 mm length preferred salmonids (rainbow trout) to cyprinids (roach) irrespective of the type of fish host, on which it had been previously grown in the laboratory. We suggest that such an innate ontogenetic shift in host preference maintains the major part of the parasite population on its principal host, ensuring successful reproduction within suitable habitats.

The free-swimming early larval stages of Argulus foliaceus (Linneaus) (Branchiura) are studied using digital video, light microscopy, and SEM. We analyze and document the mode of swimming in the hatching stage of A. foliaceus and the subsequent juvenile stages with fully developed thoracopods. We present new observations and an analysis of the functional morphology of a cleaning behavior in the first stage. This stage swims very efficiently using the large exopods of the second antennae in concert with the mandibular palp (naupliar limbs), while the subsequent stages use the now developed thoracopods for propulsion. This posterior shift in propulsion is similar to--but independent from--what is seen in other crustaceans. The hatching stage has previously been referred as a "metanauplius" but as the first and second maxillae are developed and active, and buds of all four thoracopods are present, it is too advanced to be included in the naupliar phase. The hooks of the first antennae and the distal hooks of the maxillae are demonstrated to function not only as attachment organs (to the host), but also to play a significant role in the cleaning of the naupliar swimming appendages. A digital video-based analysis of the swimming mode is provided. The larval swimming pattern is generally similar to that of other crustaceans such as Branchiopoda and Cirripedia, but autapomorphies of the Branchiura include the following: 1) While actively swimming, the naupliar appendages are almost straight during the recovery stroke and 2) they have a relatively small deflection during movement ( approximately 25 degrees or approximately 35 degrees for mandible and second antenna respectively), 3) the larval mandible has a uniramous palp which is the retained exopod. The morphological implications of the transition from the possibly nonfeeding pelagic, or free-swimming, first larval stage to the feeding, parasitic second stage are discussed and compared with other crustaceans.

In this study, metazoan parasites of bream (Abramis brama Linnaeus, 1758) in the Lake Durusu (Terkos) were investigated between June 2002 and May 2003. During this study, a total of 67 bream were examined for the presence of metazoan parasites. Ten species of parasites were found on 64 of the 67 fish examined. These parasites are: Dactylogyrus sphyrna (Linstow, 1878) and D. distinguendus (Nybelin, 1936) Monogenoidea, Caryophyllaeus laticeps (Pallas, 1781) Cestoidea, Tetracotyle sp, Diplostomum sp. and Tylodelphys clavata (Nordmann, 1832) metacercaria Trematoda, Eustrongylides excisus (Jagerskiöld, 1909) Nematoda, Piscicola geometra (Linnaeus, 1758) Hirudinea, glochidia of mollusk, Bivalvia, Argulus foliaceus (L., 1758) Crustacea. Diplostomum sp., Dactylogyrus sphyrna and D. distinguendus were found to be the dominant parasites of A. brama. Both the prevalence and intensity of other parasites were not found to be high. All identified parasites are a new finding for A. brama in the Lake Durusu. This is the first time that D. distinguendus has been identified in Turkey.

Compensatory or 'catch-up' growth following prolonged periods of food shortages is known to exist in many free-living animals. It is generally assumed that growth rates under normal circumstances are below maximum because elevated rates of growth are costly. The present paper gives experimental evidence that such compensatory growth mechanisms also exist in parasitic species. We explored the effect of periodic host unavailability on survival, infectivity and growth of the fish ectoparasite Argulus coregoni. Survival and infectivity of A. coregoni metanauplii deprived of a host for selected time periods were age dependent, which indicates that all metanauplii carry similar energy resources for host seeking. Following the periods off-host, metanauplii were allowed to settle on rainbow trout and were length measured until they reached gravidity. During early development on fish, body length of attached A. coregoni was negatively correlated with off-host period indicating a mechanism that creates size variance in an attached parasite cohort originally containing equal amounts of resources. However, over time the size differences between parasites became less pronounced and eventually parasites that were kept off-host for longest periods of time reached the length of those individuals that had been allowed to infect a host sooner. A. coregoni thus appears to compensate for delayed growth resulting from an extended host searching period by elevated growth rates, although we show that such accelerated growth incurred a cost, through decreased life-expectancy.

By sampling individual rainbow trout, Oncorhynchus mykiss, at a fish farm we showed that Argulus coregoni were aggregated within their host population. The relative significance of susceptibility and exposure generating the observed pattern was tested using experimental infections. We examined, whether rainbow trout developed protective resistance mechanisms against the louse following a challenge infection and if there was variation between individual trout in their susceptibility to A. coregoni metanauplii. Fish were exposed to 20 A. coregoni for 5, 25, 50, 85 or 120 min and the numbers attaching recorded. Three weeks later, developing argulids were removed and the experiment repeated with a standardized exposure of 20 metanauplii. Prior exposure of fish with A. coregoni did not reduce the total infection intensity compared to naive fish, but fish gained infection more rapidly. We suggest that there is no protective acquired resistance of pre-exposed rainbow trout to subsequent Argulus exposure. The possibility that an immunosuppressive mechanism by argulids was acting enabling the higher attachment rate could be refuted since control individuals, not previously exposed to lice, gained the infection at a similar rate as the fish challenged twice. Our results do not indicate clear differences in susceptibility among individual fish but the transmission of metanauplii on fish seemed to be opportunistic and non-selective. Our results support the view that variation in exposure time, rather than differences in susceptibility of individual hosts, might be the key factor in generating the aggregated distribution of Argulus on their hosts.

The obligate fish ectoparasite Argulus coregoni is strictly specific to salmonids and is very rarely found on other fish species. The ability of the parasite to grow and complete its life cycle on a cyprinid host, Rutilus rutilus, was compared with that on a typical salmonid host, Oncorhynchus mykiss. Rearing experiments were run for 42 d with newly hatched metanauplii in flow-through tanks. Body length and sex of the parasites were recorded every 5 d. Growth rates on O. mykiss exceeded those on R. rutilus from the age of 2 wk, at which time the parasites reached a length of about 3.5 mm. Males grew faster than females at the beginning of the experiment up to a length of 2.5 to 3.0 mm; thereafter, a faster growth rate was observed in females. In another experiment, association of parasites with the hosts was monitored and residence time defined as the period between attachment and first detachment from the host. Longer residence time was observed on O. mykiss than on R. rutilus; female parasites stayed on both fish species longer than did males. Faster growth of parasites could be associated with longer uninterrupted periods of attachment to hosts, since frequent detachment means higher energy losses and less time available for feeding. Despite its slower growth on R. rutilus, A. coregoni matured and laid egg clutches, but took 5 d longer than on O. mykiss. The potential of A. coregoni to complete its life cycle on cyprinids could have important ecological consequences, creating an infection reservoir when the main salmonid hosts are rare or temporarily missing.

The choice between two alternative hosts, brown trout (Salmo trutta) and roach (Rutilus rutilus), and the response to visual and olfactory cues were studied in the ontogeny of Argulus coregoni. The initial preference of the smallest parasites for brighter roach changed at the age of 2 weeks, at the size of about 2 mm, for trout, a typical salmonid host. Younger argulids were attracted by a non-specific visual stimulus (white disc over dark background), and they did not respond to olfactory stimulation (fish-conditioned water). Later, the response to visual stimuli was modulated by trout-conditioned water, but not by that conditioned by roach. The primary role of vision, particularly in early ontogeny, is emphasized as an adaptation of A. coregoni to habitats in boreal latitudes, clear and running water with a sparse fish population. In sub-adult and adult parasites, vision is complimented by olfaction that enables them to choose hosts more precisely. The nature and adaptive significance of the ontogenetic shift in host choice by A. coregoni is discussed.

The population structure and recruitment of Argulus coregoni was monitored at a Finnish fish farm during the open water periods of 1999 and 2001 by weekly sampling of attached argulids. In 2001 the numbers of rainbow trout examined increased in the autumn when the A. coregoni population was declining. When the water temperature exceeded 10 degrees C, at the end of May, A. coregoni egg hatching commenced. A mean number of 98 (S.D. +/- 5.4) juvenile A. coregoni was recorded on each fish, before the start of female egg laying in July 1999. The abundance of lice was lower in 2001. The main recruitment of A. coregoni juveniles occurred in early summer, but the hatching of eggs continued until September. However, no pulses of hatching were recorded later in the summer and the numbers of lice on fish substantially decreased after mid-July in both years. On average, only 0.6 (S.D. +/- 0.72) parasites/fish were found between August and late October 2001 and none in November. We suggest that there is 1 main A. coregoni generation annually in Central Finland. Environmental conditions, especially temperature, affects the population cycle of A. coregoni: we found a more synchronous and intense population cycle during the summer 1999, when the early summer was warmer than in 2001. The overall sex ratio (female:male) of A. coregoni was nearly 1:1 in June 1999, but was male biased (1:1.4) in June 2001. It was also shown that from July onwards, many females detached from the fish host in order to lay their eggs on the bottom of the pond. Large males were often bigger than the largest females between mid-July and early September 2001, when as many as 3.7 (s.r. +/- 1.48) times more males than females were present in the lice samples.